We have prepared plasmids containing various lengths of palindrome. When the palindrome is longer than 184bp, the plasmid can not grow in wild-type Escherichia coli, but a plasmid with a palindrome less than 95bp can. The absolute length of the palindrome determines whether the plasmid can grow or not. Electron microscopy confirmed that the plasmid with palindrome had a relaxed form with cruciform structure. The length of the protruding region is not the same size as the palindrome inserted into the plasmid but the self-annealing length seems to be just enough to make the twisted molecule into the relaxed form. Although plasmids with palindromes longer than 184bp multiply in E. coli recB recC sbcB recF, deletions occur in the palindromes and these deletions seem to occur at a specific site.A plasmid containing a long palindromic sequence has been shown not to multiply in wild-type Escherichia coli (1-4). However, a plasmid with the 73 by palindrome (half-length) derived from SV40 (pSVoriH) can be maintained in wildtype E. coli (5). LEACH and STAHL (6) found that phage ) with a 1.6 kb palindrome could multiply in E, coli recB recC sbcB, suggesting that such a palindrome is degraded by a recombination system in E. coli. In vitro, a plasmid which contains a palindromic structure becomes cruciform when treated with DNA gyrase (7). We studied a plasmid carrying two lactose promoter sequences in palindromic positions (184 by x 2) and found that it could not multiply in wild-type E. coli but could in E. coli recB recC sbcB recF. Structural analysis of the plasmid DNA by agarose gel

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We have prepared three kinds of plasmids containing two lactose promoters in palindromic positions. In the first, structure I, the promoters are joined tail to tail and transcription proceeds from head to tail. In structure II they are joined head to head. In structure III a foreign DNA is located between the two lactose promoters. The plasmid of structure I can not multiply in wild-type Escherichia coli but can in E. coli recB recC sbcB recF. Since a plasmid with structure III can grow in the wild-type strain, a linear DNA fragment which is prepared from structure III by removing the foreign DNA can be used as a very efficient cloning vector. Plasmid DNA with structures I and II isolated from E. coli recB recC sbcB recF were shown to have a relaxed form by agarose gel electrophoresis. However, in contrast to the structure I plasmid, structure II plasmid can multiply in wild-type cells, though it is unstable and present in a low copy number. Excised head to head dimer, but not tail to tail dimer, moved at an unusual rate in polyacrylamide gel electrophoresis. The difference between the physical structures of the two dimers of the lactose promoter may be reflected in the biological character of the plasmids.Several laboratories have reported that a plasmid carrying palindromic DNA longer than 73bp could not multiply in wild-type Escherichia coli (1-3). LEACH and STAHL (4) reported that phage 2 carrying a long palindrome could not multiply in wild-type E. coli but could in E. coli recB recC sbcB. MIzuuCHI et al. prepared a palindromic DNA and showed it became cruciform when it was treated by DNA gyrase in vitro (S).We have tried to construct a plasmid containing two lactose promoters in palindromic positions. The five possible structures of a plasmid with two lactose promoters are shown in Fig. 1. Structures I and II have the same lactose promoters but their direction of joining is opposite. This minor difference had a significant effect on the viability of the plasmids and their physical structure. Structure IV was

Low density lipoprotein (LDL) and high density lipoprotein (HDL) were purified from normal human serum by KBr density gradient centrifugation and gel filtration through Sepharose 4B. LDL reversibly inhibited proliferation of Swiss/ 3T3 cells, whereas HDL had no inhibitory effect on cell growth. The LDL-induced inhibition was LDL-dose dependent and was reversed by the addition of mevalonate, a product of the reaction of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase (mevalonate : NADP+ oxidore ductase (CoA-acylating), EC 1.1.1.34). These data suggest that a specific reduction in the activity of HMG-CoA reductase produced by the addition of LDL is the main cause of the inhibition of cell proliferation. Studies of the effect of LDL on the cell cycle showed that it inhibited the entry of cells arrested in G 0/G1 into the S phase but that it did not affect the transition of cells at the G1/S boundary into the M phase. The cell cycle of 3T3 is arrested solely in G1 by LDL.

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